Interleukin-4 (IL-4) and IL-13, their mutants and soluble receptors are being tested in the clinic by systemic injection or by gene transfer using viral or plasmid vectors for the treatment of rheumatoid arthritis, bronchial asthma, and cancer. Studies are underway to characterize the structure, function, signal transduction and targeting of receptors (R) for IL-4 and IL-13 on immune and cancer cells. 1) Reconstitution studies have confirmed that IL-13R alpha'chain, but not alpha chain, is a novel component of the IL-4R system. These studies have demonstrated that IL-4R and IL-13R share two chains (IL4R beta and IL-13R alpha')with each other that help explain similar biological activities of IL-4 and IL-13 on many different cell types including cancer cells. 2) To determine the subunit structure of IL-13R receptor in primary brain tumor cells reverse transcriptase-PCR (RT-PCR)analysis of RNA from primary brain tumor cells, normal astrocytes and normal brain tissue have demonstrated that transcripts of IL-13Ralpha chain is present in greater abdunance in malignant glioma cells as compared to normal astrocytes or normal brain tissues. The transcripts for two other chains (e.g., IL-13Ralpha' and IL-4Rbeta), on the other hand, yielded similar PCR positivity in brain tumors and as well as normal samples while transcripts for gamma chain were absent in all brain tumor cells and normal tissues. Indirect immunofluorescence studies for different receptor chains confirmed the RT-PCR results and demonstrated a striking difference in the level of expression of IL-13Ralpha protein between normal astrocytes and malignant astrocytoma cells. These studies establish the IL-13Ralpha subunit as a novel tumor specific protein that may be useful as a tumor marker, a target for cytotoxin/immunotoxin or alternatively, a tumor associated antigen for active, specific immunotherapy. 3) To create a mutant of IL- 13 that is cancer IL-13R selective, we created a novel mutated form of human IL-13, in which a positively charged arginine (R) at position 112 was substituted to a negatively charged aspartic acid (D). This mutant termed as IL-13R112D was expressed in E. coli and purified to near homogeneity. IL-13R112D was found to be a potent IL-13 agonist with 5-10 fold improved binding affinity to IL-13 receptors (IL-13R) compared to wild-type IL-13 (wtIL-13). The agonistic activities were revealed on the basis of ~10 fold improved activity over wtIL-13 in several assays: a) inhibition of CD14 expression in primary monocytes; b) proliferation of TF-1 and B9 cell lines; and c) activation of STAT6 in EBV-immortalized B cells, primary monocytes and THP-1 monocytic cell line. Furthermore, mutant IL-13R112D neutralized the cytotoxic activity of a fusion protein composed of wtIL-13 and a Pseudomonas Exotoxin (IL-13-PE38) approximately 10 times better than wtIL-13. Based on these results, we conclude that IL-13R112D interacts with much stronger affinity than wtIL-13 and that Arg112 plays an important role in the interaction with its receptors. Thus, these results suggest that IL-13R112D may be useful for the study of IL-13 interaction with its receptors or alternatively in designing specific targeted agents for IL-13R positive malignancies. 4) The IL-4 and IL-13R directed targeting of a Pseudomonas exotoxin, Diphtheria toxin, or alternatively receptor directed gene transfer is also being investigated. The receptors for these two interleukins are expressed in abundance on human tumor cells that offer an attractive target for toxin therapy or gene therapy. In vivo experiments in immunodefeicient mice with human breast cancer and AIDS-associated Kaposi's sarcoma tumors have demonstrated complete responses in a dose dependent manner in response to IL4-toxin and IL-13 toxin administration. Preclinical pharmacokinetics and toxicology studies with IL-13 cytotoxin has allowed us to begin a Phase I/II trial for brain tumor and renal cell carcinoma. Our previous studies on IL-4R targeting resulted in a Phase I clinical trial for malignant gliomas. Our recently published results suggest that this agent is active in brain tumor patients. A phase I clinical trial continues to enroll patients at various Centres in US and Germany. These clinical studies will help elucidate the safety and efficacy of this and other chimeric toxins being tested in clinic under various INDs.